PCR of episomal plasmids
ALASTAIR MACKAY
amackay at welchlink.welch.jhu.edu
Fri Jun 3 23:42:05 EST 1994
I agree with Brian Foley that the answer is PCR of whole cells. It seems
to me that a combination of positive controls and optimization has a good
chance of working. Positive controls: x pg or fg of plasmid doping the
whole cells, compared to this amount in absence of cells. Optimization:
Pay attention to Mg+2, do a checkerboard as Current Protocols suggests.
Also check annealing temperature. Consider adding TMAC (tetramethyl
ammonium chloride) in the 10E-5 to 10E-6 molar range, I found it can help
increase specificity.
Good luck, Alastair
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